Regulators of G protein signaling (RGS proteins) are GTPase-activating proteins (GAPs) for certain Galpha subunits and, therefore, possess the potential to regulate the active lifetimes of both Galpha and Gbetagamma subunits in vivo. A major limitation of current knowledge concerning RGS proteins is understanding their physiological roles in vivo and the role of structural differences in their signaling functions. Here we provide new evidence of extraordinary complexity in the human RGS6 protein family and the identification of novel protein interactions and signaling functions of these proteins. We identified 20 splice variant forms of RGS6, all of which possess the domain required for GAP activity toward Galpha subunits. We discovered that RGS6 protein interaction with SCG 10, a neuron-specific microtubule-destabilizing protein, promotes neuronal differentiation of PC12 cells. We also found that overexpression of RGS6 proteins in zebrafish embryos, where we identified neural expression of RGS6 transcripts, produced marked neural development effects. This proposal is designed to provide new structural and functional insights into our understanding of RGS6 proteins. The role of structural differences in RGS6 proteins and their interactions with SCG10 and Gbeta5, proteins that bind to the GGL (Ggamma-subunit like) domain of RGS6 proteins, in their ability to regulate G protein signaling and neuronal differentiation is unknown. We will characterize the role of RGS6 proteins and interactions with their binding partners on G protein signaling in cellular systems. We will define the residues or motifs within the GGL domain of RGS6 that bind G? and SCG10 and examine the specificity and generality of this effect. We will determine the mechanisms underlying the ability of RGS6 proteins to induce neurite outgrowth and microtubule disruption. We will characterize the role of RGS6 in neural development and its interaction(s) with its binding partners by gain of function and gene knockdown approaches with wild type and mutant RGS6 protein in the experimentally amenable zebrafish. This work will provide new structural and functional insights into the molecular basis of novel signaling actions of RGS proteins and will contribute in significant ways to understanding these mechanisms of RGS6 function in neural development.